There is a need in the art for sequencing the ends of long linear DNAs for various applications including, e.g., phage-display libraries and many others. Phage-displayed technology has demonstrated to be the most successful technology in producing effective therapeutic antibodies for many diseases and indications. The success of the industry depends on the quality and diversity of the phage-displayed antibody library. The diversity of the library lies in the variable regions of both the heavy chain and light chain.
In a typical phage-displayed antibody library, the VH (heavy chain variable region) and VL (light chain variable) are separated from each other by a long stretches of sequences encoding the constant regions (CH1 or CL), phage coat proteins, and secreting signals. Traditionally, Sanger sequencing were performed to sample the diversity of the library or verify the sequence of the selected clones after repeated panning. Using Sanger sequencing, both heavy chain and light chain variable region can be read but only limited number of the clones can be assessed due to the low throughput. With high throughput parallel sequencing, a vast population of the library could be read. However, with today's technology, it remains challenging to sequence both the heavy chain and light chain variable at same time. There is a need to sequence the VH and VL DNA sequence together as they are paired in the phage-display construct.